Single-cell RNA-seq analysis reveals penaeid shrimp hemocyte subpopulations and cell differentiation process

Crustacean aquaculture is expected to be a major source of fishery commodities in the near future. An immune priming system of shrimp is crucial for a sustainable supply, as shrimp do not have an adaptive immune system; however, little is known about their immunity. Hemocytes are known as key agents of the crustacean immune system; nevertheless, we have yet to identify the different cell types, functions, and differentiation and maturation processes associated with it.

To date, only discrete and inconsistent information on the classification of shrimp hemocytes has been reported, showing that the morphological characteristics are not sufficient to resolve their actual roles. Therefore, we employed a single-cell transcriptome approach for shrimp hemocytes. Thousands of hemocytes from shrimp Marsupenaeus japonicus were subjected to single-cell mRNA sequencing (scRNA-seq).

From the classification of cells based on their transcriptional profiles, we discovered nine different hemocyte subpopulations corresponding to different stages of the differentiation process that can be traced back to the first subpopulation. Using our classification, we also identified molecular markers for each subpopulation, and mapped their differentiation and maturation pathways.

Interestingly, we also discovered growth factors that may play crucial roles during the differentiation process and provide key information for hemocyte cell culture. Among these subpopulations, different immune roles were suggested from the analysis of the differentially expressed immune-related genes. The present characterization results, based on the scRNA-seq, should set the fundamental ground for understanding shrimp immunity for the future development of shrimp aquaculture.